Hyperalgesic states result from the release of primary afferent transmitter release e.g., substance P (SP) from capsaicin-sensitive C fibers which activate postsynaptic systems through an NK1 receptor. This activation leads to the spinal release of excitatory amino acids and prostaglandins (PGE2), which evokes a facilitated state through an action on NMDA and prostanoid receptors. This hormonal input leads to phosphorylation of receptors and channels which can enhance nociceptive processing. Spinal mu opioid and alpha2 adrenergic agonists produce antinociception, in part by blockade at a site presynaptic to the primary afferent leading to a block of SP release and by an action at receptors postsynaptic to the primary afferent. Continued presence of the agonists results in an increase in the dose required by a probe drug to produce a given effect. When an antagonist is delivered to a tolerant system, withdrawal is observed which is associated with hyperalgesia. Accumulating evidence suggested that these effects, injury evoked hyperalgesia, spinal tolerance and withdrawal-evoked hyperalgesia are mediated through parallel changes in spinal NMDA receptor activity and changes in the phosphorylation state of spinal neurons induced by kinases/phosphatases activated during chronic afferent input and exposure to opiates. These observations provoke several hypotheses. 1) Inhibition by mu or alpha2 agonists of capsaicin-evoked release of SP from afferent terminals or release of glutamate and PGE2 evoked by local NK1 receptor activation from nonafferent spinal terminals will be diminished by continuous exposure to mu/alpha2; 2) After chronic exposure to spinal mu or alpha2 agonists, antagonists (withdrawal) will evoke hyperalgesia and enhance release of spinal SP, PGE2/ glutamate. 3) Concurrent spinal NMDA antagonism will prevent loss of effect by mu or alpha2 agonists on evoked release and prevent release observed in antagonist-evoked withdrawal. 4) mu PKC) and phosphatases (e.g. calcineurin), which may decrease (by phosphorylation) or increase (by dephosphorylation) the ability of the agonist to reduce C transmitter release and nociception after chronic exposure. 5) Blockade of synthesis of specific kinases (PKCalpha or PKCgamma) and phosphatases (calcineurin) will reduce/enhance, respectively, tolerance and withdrawal-hyperalgesia. These studies will define i) the role of primary afferents in antinociception in the face of chronic opiate or alpha2 exposure; ii) linkages at which NMDA receptor antagonism alters spinal mu and alpha2 tolerance, and iii) the role of spinal isozymes (kinases and phosphatases) which regulate phosphorylation in the development of tolerance and hyperalgesia.